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Semen pH in patients with normal versus abnormal sperm characteristics
Semen pH in patients with normal versus abnormal sperm characteristics Carolyn Harraway, MD, Nathan G. Berger, MD, and Norman H. Dubin, PhD Baltimore, Maryland OBJECTIVE: The World Health Organization laboratory manual, last revised in 1992, states that the normal pH of semen ranges from 7.2 to 8.0. Our experience has been that values in our patient population are consistently higher than this range. To confirm this we reviewed >1100 semen records. STUDY DESIGN: All patient records from January 1994 to December 1998 that had semen pH measurements and sperm concentration and motility measurements recorded were included in this study. We also determined the semen pH in a subgroup of patients who underwent sperm preparations for intrauterine inseminations that resulted in documented pregnancies. Histograms were used to describe the populations and the Mann-Whitney test was used for group comparisons. RESULTS: For all patients (N = 1199) mean (±SD) semen pH was 8.2 ± 0.3. The range was 7.3 to 9.5, with pH <8.0 in 32% of the samples. The semen pH among the patients with normal sperm concentration and motility values (n = 602) was not different from that among those with abnormal parameters (n = 597). Mean semen pH value was 8.2 for both groups. In a small group of patients (n = 19) whose sperm preparations had been documented to result in a clinical pregnancy after intrauterine insemination the semen pH was 8.3 ± 0.3, with a range of 7.9 to 8.7. CONCLUSION: Our study questions the reference range defined by the World Health Organization for semen pH of 7.2 to 8.0. The mean values that we observed in our population, including those of samples from patients with normal sperm parameters, consistently lay outside that range. (Am J Obstet Gynecol 2000;182:1045-7.)
Key words: pH, semen, sperm, World Health Organization
The analysis of semen is routinely performed as part of the evaluation for infertility. The measurement of pH is a standard component of the semen analysis. The World Health Organization (WHO) laboratory manual, last revised in 1992, states that the normal pH of semen ranges from 7.2 to 8.0.1 Interestingly, the more recently published WHO clinical manual gives a reference range of 7.2 to 7.8.2 During the last 5 years our experience has been that semen pH in our patient population was consistently higher than the reference range. A recent article from Norway3 reported that the studied population had pH values consistently >8.0. For this reason we reviewed >1100 semen records and determined how the pH values compared with the WHO guidelines. We also compared semen pH values of patients with abnormal sperm parameters with those of patients with normal sperm parameters. Furthermore, we analyzed a subset of patients with proved fertility. From the Department of Gynecology and Obstetrics, Union Memorial Hospital. Reprints not available from the authors. Copyright © 2000 by Mosby, Inc. 0002-9378/2000 $12.00 + 0 6/1/106051 doi:10.1067/mob.2000.106051
Material and methods Before analysis of the records the reliability of the data was assessed by comparison of the accuracy and precision of various brands of pH paper (pHydrion; Micro Essentials Laboratory, Inc, Brooklyn, NY; and ColorpHast; EM Science, Gibbstown, NJ) with pH measurement by a meter (Orion model 520A; Orion Research, Inc, Beverly, Mass) for a range of standards. These brands were the ones used during the period of data collection. The meter was calibrated with pH 7 and pH 10 calibration buffers (Ricca Chemical Company, Arlington, Texas). The pH standards were prepared for a range between 7 and 8 with a monobasic potassium phosphate and sodium hydroxide buffering system.4 Precision was assessed by calculating the average coefficient of variation for replicate measurements at each standard pH. Accuracy was assessed by calculating the mean absolute difference of the measurement from the standard value. Measurements were made by one of us without knowledge of the standard pH value. Precisions (coefficients of variation) were 1.43% and 1.57% for pHydrion pH paper and ColorpHast pH paper, respectively, compared with 0.24% for the pH meter. Accuracy for the meter was within 0.004 unit of the standards; accuracies were –0.01 1045
1046 Harraway, Berger, and Dubin
Fig 1. Frequency distribution of semen pH values for our population of patients (N = 1199) compared with WHO normal limits (horizontal bar). Frequencies are expressed as absolute numbers.
May 2000 Am J Obstet Gynecol
Fig 3. Comparison of frequency distributions of semen pH from patients whose partners became pregnant after intrauterine insemination (IUI) in cycle in which pH measurement was made with that of general study population. In this and Fig 2 normal frequency distribution is inverted. Frequencies are expressed as percentages because of relatively small number of cases in intrauterine insemination group.
intrauterine inseminations that resulted in documented pregnancies. For our patient data, histograms were used to describe the populations. Because of nonnormal distribution of the data the Mann-Whitney test was used for group comparisons. Correlations of pH with other parameters were determined as noted. Results Fig 2. Comparison of frequency distributions of semen pH from patients with male factor infertility and patients with normal sperm parameters. In this and Fig 3 normal frequency distribution is inverted. Center horizontal line is 0. Frequencies are expressed as absolute numbers.
and 0.08 for the pHydrion and ColorpHast pH papers, respectively. All patient records in a computer-assisted semen analyzer database (Motion Analysis Corporation, Santa Rosa, Calif) from January 1994 to December 1998 for which semen pH measurements and sperm concentration and motility measurements were available were included in this study. All pH measurements were made with pH paper on raw semen ejaculates at the time of semen analyses, screens, or sperm preparations for intrauterine insemination and other assisted reproductive technology procedures. We determined the distribution of semen pH values for our patient population and compared values between patients with normal (concentration, >20 million cells/mL; motility, >40%) and abnormal sperm parameters. We also determined the distribution of semen pH values in a subgroup of patients who underwent sperm preparations
The mean (±SD) age of patients in our population was 36.2 ± 6.3 years. The mean (±SD) duration of abstinence before production of the specimen was 3.9 ± 3.4 days and the mean (±SD) time from specimen production to analysis was 30.2 ± 17 minutes. For all patients (n = 1199) the mean (±SD) pH was 8.2 ± 0.3, with a median of 8.2. The range was 7.3 to 9.5, with pH <8.0 in 32% of the samples. Fig 1 illustrates the frequency distribution for this patient sample and indicates the reference pH range described by WHO. The semen pH among the patients with normal sperm concentration and motility (n = 602) was not different from that among those with abnormal parameters (n = 597; Fig 2). Both means were 8.2, as were the medians. In a small group of patients (n = 19) whose sperm preparations were documented to result in a clinical pregnancy after intrauterine insemination the mean (± SD) semen pH was 8.3 ± 0.3, with a range of 7.9 to 8.7 (Fig 3). For all semen samples studied there were significant (P < .001) but relatively weak negative correlations between pH and duration of abstinence (r = –0.22), volume (r = –0.15), and concentration (r = –0.12). There was no relationship between pH and patient age or interval from collection to measurement, nor was there a change in average pH during the period studied.
Harraway, Berger, and Dubin 1047
Volume 182, Number 5 Am J Obstet Gynecol
Comment The seminal plasma is derived primarily (50%-80%) from the seminal vesicles, with a smaller fraction (13%30%) contributed by the prostate. The Cowper and Littre glands provide an additional small percentage. The pH of the ejaculate is determined predominantly by the basic seminal vesicle secretions and the prostatic secretions, which may have a pH between 6.5 and 7.2.5 With advancing age or infection this fluid may become more basic.6 The normal pH of semen has been defined as ranging from 7.2 to 8.0. Our population of >1100 patients had an average pH of 8.2, with only 32% of the specimens falling within the supposedly normal range. Haugen and Grotmol3 also noted that the semen pH in their population was consistently higher than WHO reference values. Although that group only studied 207 patients, both pH paper and a pH meter were used to analyze each sample. They found semen pH to be 8.2 with a pH meter and 8.4 with pH paper. Another group reported elevated semen pH among young healthy medical students.7 Although the pH paper used in our laboratory is not as accurate or precise as a pH meter, we demonstrated that it was accurate enough to support our findings. Furthermore, there was no correlation between pH and the date of analysis through the 5-year study period, so it is unlikely that pH was affected by changes in lots of pH paper or changes in technicians during that period. Several inflammatory processes of the prostate and seminal vesicles are thought to alter semen pH, and according to the WHO infection should be suspected if the pH exceeds 7.8.2, 6 In light of our results this recommendation could possibly lead to overdiagnosis of infection and other inflammatory processes. It is interesting that the WHO manual1 states that the optimal pH for sperm migration and survival in the cervical mucus is 7.0 to 8.5.
Although our findings confirmed those of Haugen and Grotmol,3 we also extended their findings by demonstrating that the mean semen pH among patients with normal sperm parameters was not different from that among those with abnormal sperm parameters. Furthermore, a subgroup of patients with proved fertility in the same cycle in which the pH was measured also had the same high range of semen pH. It should be noted, however, that this subgroup cannot be construed as a normal fertile population because the sperm was washed and inserted directly into the uterine cavity as part of the intrauterine insemination procedure. This finding does demonstrate, however, that exposure to high pH semen does not preclude the functional potential of sperm. Our study questions the interpretation of the reference pH range defined by WHO, which clearly does not reflect the values that we observed in our population, including those with proved functional sperm. REFERENCES
1. World Health Organization. WHO laboratory manual for the examination of human semen and sperm-cervical mucus interaction. Cambridge (UK): Cambridge University Press; 1992. 2. World Health Organization. WHO manual for the standardized investigation and diagnosis of the infertile couple. Cambridge (UK): Cambridge University Press; 1993. 3. Haugen TB, Grotmol T. pH of human semen. Int J Androl 1998;21:105-8. 4. Dawson RM, Elliot WH. Buffers and physiological media. In: Dawson RM Elliot, DC, Elliot WH, Jones KM, editors. Data for biochemical research. Oxford (UK): Oxford University Press; 1959. p. 200. 5. Partin AW, Coffey DS. The molecular biology, endocrinology, and physiology of the prostate and seminal vesicles. In: Walsh PC, editor. Volume 2: Campbell’s urology. 7th ed. Philadelphia: WB Saunders; 1998. p. 1390. 6. Meares EM Jr. Prostatitis syndromes: new perspectives about old woes. J Urol 1980;123:141-7. 7. Bhushan S, Pandey RC, Singh SP, Pandey DN, Seth P. Some observations on human semen analysis. Indian J Physiol Pharmacol 1978;22:393-6.
Key words: pH, semen, sperm, World Health Organization
The analysis of semen is routinely performed as part of the evaluation for infertility. The measurement of pH is a standard component of the semen analysis. The World Health Organization (WHO) laboratory manual, last revised in 1992, states that the normal pH of semen ranges from 7.2 to 8.0.1 Interestingly, the more recently published WHO clinical manual gives a reference range of 7.2 to 7.8.2 During the last 5 years our experience has been that semen pH in our patient population was consistently higher than the reference range. A recent article from Norway3 reported that the studied population had pH values consistently >8.0. For this reason we reviewed >1100 semen records and determined how the pH values compared with the WHO guidelines. We also compared semen pH values of patients with abnormal sperm parameters with those of patients with normal sperm parameters. Furthermore, we analyzed a subset of patients with proved fertility. From the Department of Gynecology and Obstetrics, Union Memorial Hospital. Reprints not available from the authors. Copyright © 2000 by Mosby, Inc. 0002-9378/2000 $12.00 + 0 6/1/106051 doi:10.1067/mob.2000.106051
Material and methods Before analysis of the records the reliability of the data was assessed by comparison of the accuracy and precision of various brands of pH paper (pHydrion; Micro Essentials Laboratory, Inc, Brooklyn, NY; and ColorpHast; EM Science, Gibbstown, NJ) with pH measurement by a meter (Orion model 520A; Orion Research, Inc, Beverly, Mass) for a range of standards. These brands were the ones used during the period of data collection. The meter was calibrated with pH 7 and pH 10 calibration buffers (Ricca Chemical Company, Arlington, Texas). The pH standards were prepared for a range between 7 and 8 with a monobasic potassium phosphate and sodium hydroxide buffering system.4 Precision was assessed by calculating the average coefficient of variation for replicate measurements at each standard pH. Accuracy was assessed by calculating the mean absolute difference of the measurement from the standard value. Measurements were made by one of us without knowledge of the standard pH value. Precisions (coefficients of variation) were 1.43% and 1.57% for pHydrion pH paper and ColorpHast pH paper, respectively, compared with 0.24% for the pH meter. Accuracy for the meter was within 0.004 unit of the standards; accuracies were –0.01 1045
1046 Harraway, Berger, and Dubin
Fig 1. Frequency distribution of semen pH values for our population of patients (N = 1199) compared with WHO normal limits (horizontal bar). Frequencies are expressed as absolute numbers.
May 2000 Am J Obstet Gynecol
Fig 3. Comparison of frequency distributions of semen pH from patients whose partners became pregnant after intrauterine insemination (IUI) in cycle in which pH measurement was made with that of general study population. In this and Fig 2 normal frequency distribution is inverted. Frequencies are expressed as percentages because of relatively small number of cases in intrauterine insemination group.
intrauterine inseminations that resulted in documented pregnancies. For our patient data, histograms were used to describe the populations. Because of nonnormal distribution of the data the Mann-Whitney test was used for group comparisons. Correlations of pH with other parameters were determined as noted. Results Fig 2. Comparison of frequency distributions of semen pH from patients with male factor infertility and patients with normal sperm parameters. In this and Fig 3 normal frequency distribution is inverted. Center horizontal line is 0. Frequencies are expressed as absolute numbers.
and 0.08 for the pHydrion and ColorpHast pH papers, respectively. All patient records in a computer-assisted semen analyzer database (Motion Analysis Corporation, Santa Rosa, Calif) from January 1994 to December 1998 for which semen pH measurements and sperm concentration and motility measurements were available were included in this study. All pH measurements were made with pH paper on raw semen ejaculates at the time of semen analyses, screens, or sperm preparations for intrauterine insemination and other assisted reproductive technology procedures. We determined the distribution of semen pH values for our patient population and compared values between patients with normal (concentration, >20 million cells/mL; motility, >40%) and abnormal sperm parameters. We also determined the distribution of semen pH values in a subgroup of patients who underwent sperm preparations
The mean (±SD) age of patients in our population was 36.2 ± 6.3 years. The mean (±SD) duration of abstinence before production of the specimen was 3.9 ± 3.4 days and the mean (±SD) time from specimen production to analysis was 30.2 ± 17 minutes. For all patients (n = 1199) the mean (±SD) pH was 8.2 ± 0.3, with a median of 8.2. The range was 7.3 to 9.5, with pH <8.0 in 32% of the samples. Fig 1 illustrates the frequency distribution for this patient sample and indicates the reference pH range described by WHO. The semen pH among the patients with normal sperm concentration and motility (n = 602) was not different from that among those with abnormal parameters (n = 597; Fig 2). Both means were 8.2, as were the medians. In a small group of patients (n = 19) whose sperm preparations were documented to result in a clinical pregnancy after intrauterine insemination the mean (± SD) semen pH was 8.3 ± 0.3, with a range of 7.9 to 8.7 (Fig 3). For all semen samples studied there were significant (P < .001) but relatively weak negative correlations between pH and duration of abstinence (r = –0.22), volume (r = –0.15), and concentration (r = –0.12). There was no relationship between pH and patient age or interval from collection to measurement, nor was there a change in average pH during the period studied.
Harraway, Berger, and Dubin 1047
Volume 182, Number 5 Am J Obstet Gynecol
Comment The seminal plasma is derived primarily (50%-80%) from the seminal vesicles, with a smaller fraction (13%30%) contributed by the prostate. The Cowper and Littre glands provide an additional small percentage. The pH of the ejaculate is determined predominantly by the basic seminal vesicle secretions and the prostatic secretions, which may have a pH between 6.5 and 7.2.5 With advancing age or infection this fluid may become more basic.6 The normal pH of semen has been defined as ranging from 7.2 to 8.0. Our population of >1100 patients had an average pH of 8.2, with only 32% of the specimens falling within the supposedly normal range. Haugen and Grotmol3 also noted that the semen pH in their population was consistently higher than WHO reference values. Although that group only studied 207 patients, both pH paper and a pH meter were used to analyze each sample. They found semen pH to be 8.2 with a pH meter and 8.4 with pH paper. Another group reported elevated semen pH among young healthy medical students.7 Although the pH paper used in our laboratory is not as accurate or precise as a pH meter, we demonstrated that it was accurate enough to support our findings. Furthermore, there was no correlation between pH and the date of analysis through the 5-year study period, so it is unlikely that pH was affected by changes in lots of pH paper or changes in technicians during that period. Several inflammatory processes of the prostate and seminal vesicles are thought to alter semen pH, and according to the WHO infection should be suspected if the pH exceeds 7.8.2, 6 In light of our results this recommendation could possibly lead to overdiagnosis of infection and other inflammatory processes. It is interesting that the WHO manual1 states that the optimal pH for sperm migration and survival in the cervical mucus is 7.0 to 8.5.
Although our findings confirmed those of Haugen and Grotmol,3 we also extended their findings by demonstrating that the mean semen pH among patients with normal sperm parameters was not different from that among those with abnormal sperm parameters. Furthermore, a subgroup of patients with proved fertility in the same cycle in which the pH was measured also had the same high range of semen pH. It should be noted, however, that this subgroup cannot be construed as a normal fertile population because the sperm was washed and inserted directly into the uterine cavity as part of the intrauterine insemination procedure. This finding does demonstrate, however, that exposure to high pH semen does not preclude the functional potential of sperm. Our study questions the interpretation of the reference pH range defined by WHO, which clearly does not reflect the values that we observed in our population, including those with proved functional sperm. REFERENCES
1. World Health Organization. WHO laboratory manual for the examination of human semen and sperm-cervical mucus interaction. Cambridge (UK): Cambridge University Press; 1992. 2. World Health Organization. WHO manual for the standardized investigation and diagnosis of the infertile couple. Cambridge (UK): Cambridge University Press; 1993. 3. Haugen TB, Grotmol T. pH of human semen. Int J Androl 1998;21:105-8. 4. Dawson RM, Elliot WH. Buffers and physiological media. In: Dawson RM Elliot, DC, Elliot WH, Jones KM, editors. Data for biochemical research. Oxford (UK): Oxford University Press; 1959. p. 200. 5. Partin AW, Coffey DS. The molecular biology, endocrinology, and physiology of the prostate and seminal vesicles. In: Walsh PC, editor. Volume 2: Campbell’s urology. 7th ed. Philadelphia: WB Saunders; 1998. p. 1390. 6. Meares EM Jr. Prostatitis syndromes: new perspectives about old woes. J Urol 1980;123:141-7. 7. Bhushan S, Pandey RC, Singh SP, Pandey DN, Seth P. Some observations on human semen analysis. Indian J Physiol Pharmacol 1978;22:393-6.